Lecture 14: DNA Replication & DNA Damage Flashcards
What are the 3 steps of DNA replication in E.coli?
Initiation
Elongation
Termination
What performs elongation of DNA?
Performed by DNA polymerase III
Why are primers needed in elongation?
DNA pol III requires a primer to begin synthesis of the new strand
Small RNA Primer supplies the free 3’ OH needed
What does sliding clamp do?
Keeps the DNA polymerase in close contact with DNA while it synthesises in 5’-3’ direction
How does elongation form
- DNA pol III is loaded on leading strand and synthesises leading strand from 5’-3’
- on lagging strand, uses multiple RNA primers to synthesise backwards in 5’-3’ direction
What are okazaki fragments?
Fragments of DNA on lagging strand
Why does lagging strand form a loop?
DNA pol III functions as a dimer, so loop is formed to allow polymerases to move in same direction
How are gaps between DNA and RNA primer repaired?
- repair polymerase Dpol I removed RNA primer using 5’-3’ exonixlease action and simultaneously builds new strand behind
- DNA ligase seals the gap
What polymerases are used in DNA replication?
DNA pol I: maturation of okazaki fragments
DNA pol II: main replicative polymerase
Primase: RNA polymerase, which synthesises short RNA primers
In e.coli, what is oriC?
Site where replication initiation starts, containing 2 sets of sequences
- 13-mer DNA repeats
- 9-mer DNA repeats
How is replication initiated in e.coli?
- 9-mer DNA repeats on oriC are recognised by DnaA (bound to ATP)
- this binding allows the complex to bind to 13-mer repeats as a secondary site
- slight opening of DNA allows loading of DnaB helicase
- once bound, SSBs bind to single strand regions of DNA to make sure double strand helix doesn’t form
- allows loading of entire replisome, starting with DNA primase to make primers
What is DNA gyrase?
Topoisomerase found infront of replisome to help deal with coiling changes
